Lectures 7&8- Kreb's Cycle Flashcards
1
Q
TCA Cycle
A
- Tricarboxylic acid cycle
- Just another term for the Kreb’s Cycle
2
Q
Kreb’s Cycle
A
- Main purpose is to oxidize the two carbons on pyruvate that are not already at “max oxidation” (the CH3-CO group) by removing electrons step by step to use the as reducing equivalents in the later energy extraction phase
- We are oxidizing the CH3-CO group to two CO2
- By removing these electrons, they are carried by electron carriers to the electron transport chain, where most of our ATP comes from
- Have to convert pyruvate into an acetyl group, however
- Step 1: Oxaloacetate gets condensed with the acetyl group on acetyl-CoA by citrate synthase to form citrate, and in the condensation CoA leaves. This occurs by the CH3 of the acetyl group attacking the carbonyl of oxaloacetate after getting deprotonated.
Citrate is a symmetrical molecule, with two CH2-COO- groups attached to the central carbon. The pro-S arm is always the NEW CH2-COO-. Since citrate is prochiral b/c aconitase only accepts it in a specific orientation due to its 3D active site conformation (asymmetric “landing site”), it ALWAYS adds the -OH to the pro-R arm of the citrate molecule (technically cis-aconitate) to form isocitrate. Since it is the pro-R arm that undergoes this, it is the OLD CH2-COO- that is being modified in the rest of the Krebs cycle????? This is a very exergonic rxn, so it is thus a committed step
- Step 2a: Citrate undergoes a dehydration via the enzyme aconitase, turning it into cis-aconitate
- Step 2b: Hydration occurs, also via aconitase, to turn cis-aconitate into isocitrate
- Step 2 Overall: citrate gets converted into isocitrate.
- Step 3: Isocitrate gets decarboxylated via isocitrate dehydrogenase to form alpha-ketoglutarate
- Step 4: alpha-ketoglutarate then gets decarboxylated via the alpha-ketoglutarate dehydrogenase complex, and instead of a proton coming in, CoA comes in, attaching to form succinyl-CoA. The alpha-ketoglutarate complex’s mechanism is practically identical to the pyruvate dehydrogenase complex
- Step 5: succinyl-CoA gets phosphorylated by a free Pi floating around by succinyl-CoA synthetase, which releases CoA. Then the same enzyme takes that Pi off of succinyl-CoA and adds it to GDP to form GTP, giving us succinate
- Step 6: Succinate gets oxidized by FAD and succinate dehydrogenase (which is bound very tightly to the mitochondria), giving off FADH2 and giving us fumarate
- Step 7: Fumarate gets hydrated with the help of fumarase to give L-Malate
- Step 8: The hydroxyl group of L-Malate gets oxidized to the keto form of the molecule, oxaloacetate, by NAD+ and malate dehydrogenase. This reaction has a very large ∆G°’, but is able to go forward spontaneously because in the Krebs cycle, the oxaloacetate is getting removed to be used in the cycle again, so the ∆G is actually negative since we are not at standard concentrations
3
Q
acetyl group
A
- A carbonyl with a methyl group attached to the carbonyl carbon
- It is oxidized in the Kreb’s cycle to give electrons that can be used in the electron transport chain
- This acetyl group has many uses in metabolism, such as a fuel source, a source of carbon for anabolism, a tag added to proteins to alter their function, etc
- Many molecules are broken down into acetate, and many other molecules are constructed from acetate, so
4
Q
CoA
A
- Coenzyme A; a coenzyme (obviously)
- Is a coenzyme to a lot of different enzymes, since it carries acetyl groups around to different enzymes
- It’s “business end” is a thiol, which attaches to the acetyl group formed from pyruvate to form acetyl-CoA
5
Q
CoA-SH
A
- Acetyl CoA but showcasing its thiol
6
Q
HS-CoA
A
- Still acetyl CoA showcasing its thiol
7
Q
Thioester bond
A
- The bond between thiol and a carbonyl. In our cases, it will be a bond between the thiol on CoA and the carbonyl of the acetyl group CoA is carrying
8
Q
Iron sulfur cluster
A
G
9
Q
NAD/NADH
A
- Electron carriers and cofactors. Are present in the conversion of pyruvate to an acetyl group as well as in the Kreb’s cycle and many other metabolic pathways
10
Q
FAD/FADH2
A
- A cofactor
- Unlike NAD, it is bound to the enzyme it is a cofactor to, so it cannot move around and carry electrons places. Is a cofactor to multiple enzymes in the conversion of pyruvate to an acetyl group as well as in the Kreb’s cycle and many other metabolic pathways
- Its business end is a isoalloxazine ring
11
Q
Pyruvate dehydrogenase complex
A
- An enzyme that essentially is made up of three smaller (but still big) enzymes, each of which have their own subunits, that catalyze different reactions
- It decarboxylates pyruvate to an acetyl group, and then attaches the acetyl group to CoA to form acetyl CoA
- Uses NAD+
- Since we are forming CO2, this is a committed step, basically, since you can’t just contain the CO2 to go backwards to form pyruvate again
- The three enzymes are, in order : pyruvate dehydrogenase, dihydrolipoyl transacetylase, and dihydrolipoyl dehydrogenase
12
Q
TPP cofactor
A
- The cofactor to pyruvate dehydrogenase, the first enzyme in the pyruvate dehydrogenase complex
- Is a prothetic group
- Its “business end” is a thiazolium ring, which can become deprotonated to give a carbanion.
13
Q
Lipoic acid cofactor
A
- The cofactor for Dihyrolipyl Transacetylase, the second enzyme in the pyruvate dehydrogenase complex
- Is a prosthetic group attached to a lysine residue coming off of the dihydrolipyl transacetlyase enzyme
- Referred to as the “lipoyl-lysine arm”
14
Q
FAD Cofactor
A
- One of the cofactors of Dihydrolipoyl Dehydrogenase
- Is a prothetic group, so it can’t leave the enzyme and go to different enzymes/molecules like NAD+ can
15
Q
Prochiral
A
- A molecule that contains a site that is not itself chiral, but can be made chiral through the addition of another moiety
- In other words, a molecule that can go from achiral to chiral in a single step
- Citrate is an example
- Citrate itself is achiral, but when aconitase comes in, it acts/becomes chiral, even though nothing about citrate itself changed, b/c aconitase has a specific “landing site” where citrate can only bind one particular way
16
Q
Amphibolic
A
- A cycle that does both catabolism and anabolism
- The Krebs cycle is an example of an amphibolic pathway, since while it is clearly catabolic, as seen in our exploration, it is also anabolic b/c the different molecules that are produced in the pathway (CIA Secret Service Fucked Me Over) can be used in other pathways that are anabolic.
17
Q
Anapleurotic reactions
A
- Reactions that replenish the intermediates in the Krebs cycle that are lost to other pathways that are anabolic
- These reactions are typically reactions that take products from glycolysis and turn them into intermediates in the Krebs cycle
- They also typically require ATP
18
Q
Pyruvate decarboxylase
A
- Another name for pyruvate dehydrogenase?
19
Q
Biotin
A
- A cofactor to pyruvate carboxylase in the production of oxaloacetate from pyruvate
- First, bicarbonate dephosphorylates ATP, giving a carboxyphosphate intermediate, which gets decarboxylated to give off CO2
- This CO2 then interacts with biotin and becomes covalently bound, giving a carboxy-biotinyl enzyme
- Then this carboxy-biotinyl complex gets decarboxylated, forming an enol state
- Pyruvate then comes in and biotin deprotonates it while coming back to its keto form, forming an enol from pyruvate
- This enol form of the pyruvate then attacks carbon dioxide to get to its keto form, giving us oxaloacetate
20
Q
Glyoxylate cycle
A
G
21
Q
Glyoxosome
A
H
22
Q
Mitochondria outer membrane
A
- Small molecules can pass though fairly easily through pores called mitochondrial porin in the outer membrane
23
Q
Mitochondrial porin
A
- The pores in the outer membrane that small molecules can pass through fairly easily
24
Q
Mitochondrial inner membrane
A
- Very impermeable and very regulated
25
Mitochondrial matrix
- Inside the inner membrane where most of cellular respiration takes place
26
Cellular Respiration
- Composed of glycolysis, the Kreb's cycle, and oxidative phosphorylation? Lectures slides seem to make it seem like it is only the Kreb's cycle and oxidative phosphorylation
- Used to create ATP from glucose to allow the cell to do work
- Requires oxygen
- Occurs in the mitochondria
27
e- transport chain
- Where we get most of our ATP from as electrons are exergonically moved down the e- transport chain
- O2 is our final electron acceptor, and is where oxygen finally comes into play in cellular respiration
28
MPC
Mitochondrial pyruvate carrier
| - Carries pyruvate into the matrix
29
Post-glycolysis Pre-Kreb's cycle steps
- Step 1: Pyruvate is in the cytosol after glycolysis. It gets inside the mitochondrial outer membrane easily, and then gets transported into the matrix via the MPC
- Step 2: Pyruvate gets decarboxylated to an acetyl group via the pyruvate decarboxylase and is attached to CoA to form acetyl-CoA. This is catalyzed by the pyruvate dehydrogenase complex
30
Pyruvate dehydrogenase
- The first enzyme of the pyruvate dehydrogenase complex
- Its coenzyme is TPP (thiamine pyrophosphate)
- The TPP can get deprotonated by another group on the enzyme, giving a carbanion that attacks the carbonyl of pyruvate that is attached to the methyl group, forming an intermediate (hydroxyethyl TPP) in which the pyruvate is attached to the TPP???
- The proton that ends up on the decarboxylated pyruvate comes from solution
31
Dihydrolipoyl Transacetylase
- The second enzyme in the pyruvate dehydrogenase complex
- Its cofactors are Lipoic acid, which forms a lipoyl-lyisne arm, and coenzyme A
- Only catalytic activity, really, is to transfer the acetyl group from the lipoyl-lysine arm to CoA
- It oxides the hydroxyethyl TPP ____ intermediate to an acetyl group, and thus gets reduced to the lipoyllysine arm's reduced, open form, where the acetyl group is attached to one of is thiols. It then transfers the acetyl group to CoA.
32
Lipoyl-lyisne arm
- Composed of the lipoid acid cofactor and the lysine residue of Dihydolipoyl transacetylase that is is attached to (31:15 of Lecture 7)
- It can "move around" (still connected to enzyme 2) to all the active sites to connect the three active sites for the three catalytic activities
33
Dihydrolipoyl Dehydrogenase
- Has FAD and NAD as its coenzymes
- FAD is a prothetic group, so it can't leave the enzyme, but NAD can
- Its purpose is to restore the enzyme back to its original structure
- Its FAD coenzyme oxidizes the lipoyllysine back to its "normal," oxidized state, and then, now as FADH2, it gets reduced back to FAD by NAD, forming NADH, and restoring the enzyme
34
NADF
- A coenzyme to dihydrolipoyl dehydrogenase?
35
pyruvate carboxylase
- An enzyme that, with the help of ATP, can turn pyruvate and HCO3- into oxaloacetate to replenish the Krebs cycle when oxaloacetate is removed for use in anabolic reactions
- Requires ATP
36
Regulated Steps in Krebs Cycle
- Pyruvate ---> acetyl-CoA (technically before krebs)
- Acetyl CoA+ oxaloacetate------> citrate (Step 1)
- isocitrate-----> alpha-ketoglutarate (Step 3)
- Alpha-ketoglutarate -------> succinyl-CoA (Step 4)
37
At which steps in the Kreb's cycle is NAD+ used?
Step 3: Isocitrate ----> alpha-ketoglutarate via isocitrate dehydrogenase
Step 4: Alpha-ketoglutarate -----> succinyl-CoA via alpha-ketoglutarate dehydrogenase complex
Step 8: Malate -------> oxaloacetate via malate dehydrogenase
38
Write out the net reaction of PDH
done
39
Draw out the structure of oxaloacetate
done
